Novel hydrophilic derivatives of 2-selenophene-4-quinolones as anticancer agents

ABSTRACT

2-aryl-4-quinolones are converted into phosphates by reacting with tetrabenzyl pyrophosphate to form dibenzyl phosphates thereof, which are then subject to hydrogenation to replace dibenzyl groups with H, followed by reacting with Amberlite IR-120 (Na +  form) to form disodium salts. The results of preliminary screening revealed that these phosphates showed significant anti-cancer activity. A novel intermediate, 2-selenophene 4-quinolone and N,N-dialkylaminoalkyl derivatives of 2-phenyl-4-quinolones are also synthesized. These novel intermediates exhibited significant anticancer activities.

FIELD OF THE INVENTION

The present invention relates to novel phosphate derivatives of2-aryl-4-quinolones, and novel intermediates, 2-selenophene 4-quinolonesand N,N-dialkylaminoalkyl derivatives of 2-phenyl-4-quinolones; and inparticular to their uses in treating human cancers.

BACKGROUND OF THE INVENTION

Quinolone derivatives were initially discovered as the agents to act onbacterial DNA gyrase, and thus developed as anti-bacterial agents.Recently DNA topoisomerase II has emerged as the pharmacological targetfor this class of quinolone compounds. We have synthesized a series ofsubstituted 2-phenyl-4-quinolone (A) which appeared to function as novelantimitotic agents. [Kuo, S. C., Lee, H. Z., Juang, J. P., Lin, Y. T.,Wu, T. S., Chang, J. J., Lednicer, D., Paull, K. D., Lin, C. M., Hamel,E. Synthesis and cytotoxicity of 1,6,7,8-substituted 2-(4′-substitutedphenyl)-4-quinolones and related compounds: identification asantimitotic agents interacting with tubulin. J. Med. Chem. 1993, 36,1146-56; Li, L., Wang, H. K., Kuo, S. C., Wu, T. S., Mauger, A., Lin. C.M., Hamel, E. Lee, K. H. Antitumor agents. 155. Synthesis and biologicalevaluation of 3′,6,7-substituted 2-phenyl-4-quinolones asantimicrotubule agents. J. Med. Chem. 1994, 37, 3400-7] Later on wecontinued to synthesize many related analogs such as2-phenylnaphthyridine-4-ones (B) [Chen, K., Kuo, S. C., Hsieh, M. C.,Mauger, S A., Lin, C. M., Hamel, E., Lee, K. H. Antitumor agents. 174.2′,3′,4′,5,6,7-Substituted 2-phenyl-1,8-naphthyridin-4-ones: theirsynthesis, cytotoxicity, and inhibition of tubulin polymerization. J.Med. Chem. 1997, 40, 2266-75], 2-phenyl-4-quinazolones (C) [Xia, Y.,Yang, Z. Y., Hour, M. J., Kuo, S. C., Xia, P., Bastow, K. F., Nakanishi,Y, Namrpoothiri, P., Hackl, T., Hamel, E., Lee, K. H. Antitumor Agents.Part 204: Synthesis and Biological Evaluation of Substituted 2-ArylQuinazolinones, Bioorg. Med. Chem. Lett. 2001, 11, 1193-6; Hour, M. J.,Huang, L. J., Kuo, S. C., Xia, Y., Bastow, K. F., Nakanishi, Y., Hamel,E., Lee, K. H. 6-Alkylamino- and2,3-dihydro-3′-methoxy-2-phenyl-4-quinazolinones and related compounds:their synthesis, cytotoxicity, and inhibition of tubulin polymerization.J. Med. Chem. 2000, 43, 4479-87] and tetrahydro-2-phenyl-4-quinolones(D) [Xia, Y., Yang, Z. Y., Xia, P., Bastow, K. F., Tachibana, Y., Kuo,S. C., Hamel, E., Hackl. T., Lee, K. H. Antitumor agents. 181. Synthesisand biological evaluation of6,7,2′,3′,4′-substituted-1,2,3,4-tetrahydro-2-phenyl-4-quinolones as anew class of antimitotic antitumor agents. J. Med. Chem. 1998, 41.1155-62], which enable us to establish structure and activityrelationships (SAR). Among these analogs, we have discovered quite a fewcompounds possessing potent cytotoxicity, such as 3′,6-disubstituted2-phenyl-4-quinolones (A-1) etc [Li, L., Wang, H. K., Kuo, S. C., Wu, T.S., Lednicer, D., Lin, C. M., Hamel, E., Lee, K. H. Antitumor agents.150. 2′,3′,4′,5′,5,6,7-substituted 2-phenyl-4-quinolones and relatedcompounds: their synthesis, cytotoxicity, and inhibition of tubulinpolymerization. J. Med. Chem. 1994, 37, 1126-35]. However, most of thecompounds with potent cytotoxicity were very lipophilic, and therefore,not suitable for in vivo and clinical studies. We thus made attempt tosynthesize hydrophilic derivatives of these 2-aryl-4-quinolone skeletonsin order to improve pharmacokinetic properties suitable for in vivo andclinical studies.

SUMMARY OF THE INVENTION

Preferred embodiments of the present invention include (but not limitedthereto) the following items:

1. A phosphate derivative of 2-aryl-4-quinolone having the followingformulas Ia, Ib or Ic:

wherein

R₂′, R₃′, R₄′, R₅′ and R₆′ independently are H, (CH₂)_(n)CH₃,(CH₂)_(n)YH, Y(CH₂)_(n)CH₃, Y(CH₂)_(n)YH, Y(CH₂)_(n)NR₈R₉, X,(CH₂)_(n)NR₈R₉,

wherein n is an integer of 0-4, Y is O or S, X is F, Cl, or Br, and R₈and R₉ independently are H, (CH₂)_(n)YH,(CH₂)_(n)N(C_(n)H_(2n+1))(C_(m)H_(2m+1)) or (CH₂)_(n)CH₃, wherein n andY are defined as above, and m is an integer of 0-4;

R₂, R₃, R₄ and R₅ independently are H, (CH₂)_(n)CH₃, (CH₂)_(n)YH,Y(CH₂)_(n)CH₃, Y(CH₂)_(n)YH, Y(CH₂)_(n)NR₈R₉, X, (CH₂)_(n)NR₈R₉,

or R₃ and R₄ together is —Y(CH₂)_(n)Y—, wherein n, Y, X, R₈ and R₉ aredefined as above; and

R₁ and R₁′ independently are H, Li⁺, Na⁺, K⁺, N⁺R₈R₉R₁₀R₁₁ or benzylwherein R₁₀ and R₁₁ independently are H, (CH₂)_(n)YH,(CH₂)_(n)N(C_(n)H_(2m+1))(C_(m)H_(2m+1)) or (CH₂)_(n)CH₃, n, m, R₈ andR₉ are defined as above.

2. The phosphate derivative according to Item 1, which has the formulaIa.

3. The phosphate derivative according to Item 2, wherein R₂′, R₃′, R₄′,R₅′ and R₆′ are all H; or one of R₂′, R₃′, R₄′, R₅′ and R₆′ is F, OCH₃or (CH₂)_(n)NR₈R₉, and the others thereof are H, wherein n, R₈ and R₉are defined as in Item 1.

4. The phosphate derivative according to Item 2, wherein R₂, R₃, R₄, andR₅ are all H; or one of R₂, R₃, R₄, and R₅ is F, OCH₃, Y(CH₂)_(n)CH₃ or(CH₂)_(n)NR₈R₉, and the others thereof are H; or R₂ and R₅ are H, and R₃and R₄ together is —O(CH₂)_(n)O—, wherein n, Y, R₈ and R₉ are defined asin Item 1.

5. The phosphate derivative according to Item 2, wherein R₁ and R₁′ areboth H or both Na⁺.

6. The phosphate derivative according to Item 5, wherein R₂ and R₅ areH, and R₃ and R₄ together is —O(CH₂)O—; and R₂′, R₃′, R₄′ and R₅′ areall H, and R₆′ is F.

7. The phosphate derivative according to Item 5, wherein R₂ and R₅ areH, and R₃ and R₄ together is —O(CH₂)O—; and R₂′, R₃′, R₄′ and R₆′ areall H, and R₅′ is F.

8. The phosphate derivative according to Item 5, wherein R₄ is F, andR₂, R₃ and R₅ are H; and R₂′, R₃′, R₄′, R₅′ and R₆′ are all H.

9. The phosphate derivative according to Item 5, wherein R₂, R₃, R₄ andR₅ are all H; and R₂′, R₃′, R₄′, R₅′ and R₆′ are all H.

10. The phosphate derivative according to Item 5, wherein R₄ is OCH₃,and R₂, R₃ and R₅ are H; and R₅′ is F, and R₂′, R₃′, R₄′ and R₆′ are H.

11. The phosphate derivative according to Item 5, wherein R₂ and R₅ areH, and R₃ and R₄ together is —O(CH₂)O—; and R₂′, R₃′, R₄′ and R₆′ areall H, and R₅′ is OCH₃.

12. The phosphate derivative according to Item 5, wherein R₄ isCH₂N(C₂H₅)₂, and R₂, R₃ and R₅ are H; and R₆′ is F, and R₂′, R₃′, R₄′and R₅′ are H.

13. The phosphate derivative according to Item 5, wherein R₄ isCH₂N(C₂H₅)₂, and R₂, R₃ and R₅ are H; and R₂′, R₃′, R₄′, R₅′ and R₆′ areall H.

14. The phosphate derivative according to Item 5, wherein R₄ is OCH₃,and R₂, R₃ and R₅ are H; and R₅′ is CH₂N(C₂H₅)₂, and R₂′, R₃′, R₄′ andR₆′ are H.

15. The phosphate derivative according to Item 1, which has the formulaIb.

16. The phosphate derivative according to Item 15, wherein R₂, R₃, R₄,and R₅ are all H; or one of R₂, R₃, R₄ and R₅ is F or OCH₃, and theothers thereof are H; or R₂ and R₅ are H, and R₃ and R₄ together is—O(CH₂)_(n)O—, wherein n is defined as in Item 1.

17. The phosphate derivative according to Item 15, wherein R₂′, R₃′ andR₄′ are all H; or one of R₂′, R₃′ and R₄′ is F or OCH₃, and the othersthereof are H.

18. The phosphate derivative according to Item 15, wherein R₁ and R₁′are benzyl.

19. The phosphate derivative according to Item 18, wherein R₂′, R₃′,R₄′, R₂ and R₅ are all H, and R₃ and R₄ together is —O(CH₂)O—.

20. A pharmaceutical composition for the killing of solid cancer cells,which comprises a therapeutically effective amount of a phosphatederivative of 2-aryl-4-quinolone as set forth in any one of Item 1 toItem 19 or a pharmaceutically acceptable salt thereof, as an activeingredient, in admixture with a pharmaceutically acceptable carrier ordiluent for the active ingredient, wherein the solid cancer cellscomprise human breast cancer, colon cancer, lung cancer, melanoma,ovarian cancer, renal cancer, stomach cancer, prostate cancer, ileocecalcarcinoma, glioblastoma, bone cancer, epidermoid carcinoma of thenasopharynx, hepatoma or leukemia cancer.

21. The pharmaceutical composition according to Item 20, wherein thesolid cancer cells are human breast cancer, colon cancer, lung cancer,renal cancer, hepatoma, or leukemia cancer

22. The pharmaceutical composition according to Item 21, wherein thesolid cancer cells are human breast cancer or colon cancer.

23. A compound of 2-selenophene 4-quinolone having the followingformulas IIb or IIc:

wherein

R₂′, R₃′ and R₄′ independently are H, (CH₂)_(n)CH₃, (CH₂)_(n)YH,Y(CH₂)_(n)CH₃, Y(CH₂)_(n)YH, Y(CH₂)_(n)NR₈R₉, X, or (CH₂)_(n)NR₈R₉,wherein n is an integer of 0-4, Y is O or S, X is F, Cl, or Br, and R₅and R₉ independently are H, (CH₂)_(n)YH,(CH₂)_(n)N(C_(n)H_(2n+1))(C_(m)H_(2m+1)) or (CH₂)_(n)CH₃, wherein n andY are defined as above, and m is an integer of 0-4;

R₂, R₃, R₄ and R₅ independently are H, (CH₂)_(n)CH₃, (CH₂)_(n)YH,Y(CH₂)_(n)CH₃, Y(CH₂)_(n)YH, Y(CH₂)_(n)NR₈R₉, X, (CH₂)_(n)NR₈R₉,

or R₃ and R₄ together is —Y(CH₂)_(n)Y—, wherein n, Y, X, R₈ and R₉ aredefined as above.

24. The compound according to Item 23, wherein R₂, R₃, R₄, and R₅ areall H; or one of R₂, R₃, R₄ and R₅ is F or OCH₃, and the others thereofare H; or R₂ and R₅ are H, and R₃ and R₄ together is —O(CH₂)_(n)O—,wherein n is defined as in Item 19.

25. The compound according to Item 24, wherein R₂′, R₃′ and R₄′ are allH; or one of R₂′, R₃′ and R₄′ is F or OCH₃, and the others thereof areH.

26. The compound according to Item 23 which has the formula IIb.

27. The compound according to Item 26, wherein R₂′, R₃′, R₄′, R₂ and R₅are all H, and R₃ and R₄ together is —O(CH₂)O—.

28. A pharmaceutical composition for the killing of solid cancer cells,which comprises a therapeutically effective amount of a compound of2-selenophene 4-quinolone as set forth in any one of item 23 to Item 27or a pharmaceutically acceptable salt thereof, as an active ingredient,in admixture with a pharmaceutically acceptable carrier or diluent forthe active ingredient, wherein the solid cancer cells comprise humanbreast cancer, colon cancer, lung cancer, melanoma, ovarian cancer,renal cancer, stomach cancer, prostate cancer, ileocecal carcinoma,glioblastoma, bone cancer, epidermoid carcinoma of the nasopharynx,hepatoma or leukemia cancer.

29. The pharmaceutical composition according to Item 28, wherein thesolid cancer cells are human breast cancer, colon cancer, lung cancer,renal cancer, hepatoma, or leukemia cancer.

30. A compound of 2-phenyl-4-quinolone having the following formula IIa:

wherein

R₂′, R₃′, R₄′, R₅′ and R₆′ independently are H, (CH₂)_(n)CH₃,(CH₂)_(n)YH, Y(CH₂)_(n)CH₃, Y(CH₂)_(n)YH, Y(CH₂)_(n)NR₈R₉, X,(CH₂)_(n)NR₈R₉,

wherein n is an integer of 0-4, Y is O or S, X is F, Cl, or Br, and R₈and R₉ independently are H, (CH₂)_(n)YH,(CH₂)_(n)N(C_(n)H_(2n+1))(C_(m)H_(2m+1)) or (CH₂)_(n)CH₃, wherein n andY are defined as above, and m is an integer of 0-4;

R₂, R₃, R₄ and R₅ independently are H, (CH₂)_(n)CH₃, (CH₂)_(n)YH,Y(CH₂)_(n)CH₃, Y(CH₂)_(n)YH, Y(CH₂)_(n)NR₃R₉, X, (CH₂)_(n)NR₈R₉,

or R₃ and R₄ together is —Y(CH₂)_(n)Y—, wherein n, Y, X, R₈ and R₉ aredefined as above;

provided that one of R₂, R₃, R₄ and R₅ is (CH₂)_(q)NR₈R₉, or one of R₂′,R₃′, R₄′, R₅′ and R₆′ is (CH₂)_(q)NR₈R₉, wherein q is an integer of I-4,and R₈ and R₉ are defined as above.

31. The compound according to Item 30, wherein R₄ is CH₂)_(q)NR₈R₉, andR₂, R₃ and R₅ are H, wherein q, R₈ and R₉ are defined as in Item 30.

32. The compound according to Item 30, wherein R₅′ is CH₂)_(q)NR₈R₉, andR₂′, R₃′, R₄′ and R₆′ are H, wherein q, R₈ and R₉ are defined as in Item30.

33. The compound according to Item 31, wherein R₄ is CH₂N(C₂H₅)₂, R₆′ isF, and R₂′, R₃′, R₄′ and R₅′ are H.

34. The compound according to Item 31, wherein R₄ is CH₂N(C₂H₅)₂, R₂′,R₃′, R₄′, R₅′ and R₆′ are all H.

35. The compound according to Item 32, wherein R₄ is OCH₃, and R₂, R₃and R₅ are H; and R₅′ is CH₂N(C₂H₅)₂, and R₂′, R₃′, R₄′ and R₆′ are H.

36. A pharmaceutical composition for the killing of solid cancer cells,which comprises a therapeutically effective amount of a compound of2-phenyl 4-quinolone as set forth in any one of Item 30 to Item 35 or apharmaceutically acceptable salt thereof, as an active ingredient, inadmixture with a pharmaceutically acceptable carrier or diluent for theactive ingredient, wherein the solid cancer cells comprise human breastcancer, colon cancer, lung cancer, melanoma, ovarian cancer, renalcancer, stomach cancer, prostate cancer, ileocecal carcinoma,glioblastoma, bone cancer, epidermoid carcinoma of the nasopharynx,hepatoma or leukemia cancer.

37. The pharmaceutical composition according to Item 36, wherein thesolid cancer cells are leukemia cancer.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows effects of compound I-1 and compound I-1-b on MCF7 tumorgrowth in a mouse xenograft model. Female SCID mice received injectionsof MCF7 transfectants to induce tumor xenografts. Mice were divided intofive groups. The second to fifth groups were given i.p. with compoundsI-1 (15 mg/kg), I-1 (30 mg/kg), I-1-b (22.5 mg/kg), and I-1-b (45mg/kg), respectively, three times per week. Data are expressed as meanof tumor weights (g)±S.E.M.*p<0.05 compared with the control.

FIG. 2 shows effect of compound I-1-b on animal survival. BALB/c micewere intraperitoneally injected with CT-26 tumor cells for 7 days beforebeginning the treatments with compound I-1-b (5 mg/kg/day and 10mg/kg/day QD×7).

FIG. 3 shows effect of quinolone derivatives on the viability of humanbreast cancer cells. MCF7 cells were treated with DMSO (Control) orvarious concentrations (0.125 μM to 10 μM) of quinolone derivative for48 hours and subsequent cell viability was measured by MTT assay.Results from three separate experiments were averaged and are presentedas mean±standard error as shown.

DETAILED DESCRIPTION OF THE INVENTION

As shown in the following Examples 1 to 6, when 2-phenyl-4-quinolones(I-1 to I-6) was reacted with tetrabenzyl pyrophosphate in the presenceof alkali, the corresponding phosphoric acid dibenzyl esters (I-1-a toI-6-a) were obtained. Catalytic hydrogenation of compounds (I-1-a toI-6-a) in alcohol affords the corresponding phosphoric acid mono esters(I-1-b to I-6-b), which could be led to water soluble salts (I-1-c toI-6-c).

Example 1

Dibenzyl 2-(2′-fluorophenyl)-6,7-methylenedioxyquinolin-4-yl-phosphate(I-1-a)

Sodium hydride (13.7 mg, 0.57 mmol) was added at 0° C. to a stirredsolution of compound I-1 (64.5 mg, 0.23 mmol) in dry tetrahydrofuran (10ml). After 1 h, tetrabenzyl pyrophosphate (100 mg, 0.19 mmol) was addedand the stirring was continued for 20 min.

The mixture was filtered, and the filtrate was concentrated under vacuumat a temperature below 35° C. The residue was dissolved indichloromethane, washed with an aqueous solution of sodium hydrogencarbonate, dried over MgSO₄ and concentrated under vacuum to givecompound I-1-a (69.1 mg, 67%)

MP 101-104° C.

¹H-NMR (CDCl₃, 300 MHz): δ 8.01-8.02 (m, 1H, H-5′), 7.77 (s, 1H, H-5),7.16-7.43 (m, 14H, H-3, H-3′, H-4′, H-6′, Ph), 7.05 (s, 1H, H-8), 6.12(s, 2H, OCH₂O), 5.26 (s, 2H, —CH₂ -Ph), 5.20 (s, 2H, —CH₂ -Ph)

MS (m/z) 544 (ES+)

Anal. calcd for C₃₀H₂₅FNO₆P: C, 66.30; H, 4.27; N, 2.58. Found: C,66.28; H, 4.35; N, 2.55.

2-(2′-Fluorophenyl)-6,7-methylenedioxyquinolin-4-yl-phosphate (I-1-b)

A suspension of compound I-1-a (97.7 mg, 0.18 mmol) in anhydrous MeOH(10 ml) was submitted to hydrogenation in the presence of 10% Pd/C (50mg) at room temperature for 10 min. The catalyst and precipitates wascollected and dissolved in 10% NaHCO₃ solution then filtered. Thefiltrate was acidified with dil HCl, the solid was then collected byfiltration and washed with acetone to give compound I-1-b (63.5 mg,97.2%).

MP>300° C.

¹H-NMR (DMSO-d6, 300 MHz): δ 7.93-7.98 (m, 1H, H-5′), 7.74 (s, 1H, H-5),7.49-7.54 (m, 1H, H-4′), 7.32-7.41 (m, 4H, H-3, H-8, H-3′, H-6′), 6.22(s, 2H, OCH₂O).

MS (m/z) 362 (ES−)

Anal. calcd for C₁₆H₁₃FNO₆P: C, 52.91; H, 3.05; N, 3.86. Found: C,52.73; H, 3.10; N, 3.81.

Sodium 2-(2′-fluorophenyl)-6,7-methylenedioxyquinolin-4-yl-phosphate(I-1-c)

Compound I-1-b was added to a mixture of 20 ml Amberlite IR-120 (Na⁺form) and 20 ml water, and then stirred for 6 h at room temperature. Themixture was then filtered to remove Amberlite, and then lyophilized togive I-1-c (49.1 mg, 69%).

¹H-NMR (D2O, 200 MHz): δ 7.48-7.66 (m, 2H, H-4′, H-6′), 7.40 (s, 1H,H-8), 7.31-7.35 (m, 1H, H-5), 7.11-7.19 (m, 2H, H-3′, H-5′), 7.03 (s,1H, H-3), 5.92 (s, 2H, OCH₂O).

Example 2

Dibenzyl 2-(3′-fluorophenyl)-6,7-methylenedioxyquinolin-4-yl-phosphate(I-2-a)

Sodium hydride (13.7 mg, 0.57 mmol) was added at 0° C. to a stirredsolution of compound I-2 (64.5 mg, 0.23 mmol) in dry tetrahydrofuran (10ml). After 1 h, tetrabenzyl pyrophosphate (100 mg, 0.19 mmol) was addedand the stirring was continued for 20 min.

The mixture was filtered, and the filtrate was concentrated under vacuumat a temperature below 35° C. The residue was dissolved indichloromethane, washed with an aqueous solution of sodium hydrogencarbonate, dried over MgSO₄ and concentrated under vacuum to givecompound I-2-a (85.6 mg, 83%).

MP 94-96° C.

¹H-NMR (DMSO-d6, 200 MHz): δ 7.61-7.78 (m, 2H, H-2′, H-4′), 7.48-7.56(m, 1H, H-5′), 7.24-7.45 (m, 13H, H-5, H-8, H-6′, Ph), 7.10 (s, 1H,H-3), 6.21 (s, 2H, OCH₂O), 5.29 (s, 2H, —CH₂ -Ph), 5.24 (s, 2H, —CH₂-Ph)

MS (m/z) 544 (ES+)

Anal. calcd for C₃₀H₂₅FNO₆P: C, 66.30; H, 4.27; N, 2.58. Found: C,66.25; H, 4.34; N, 2.55.

2-(3′-Fluorophenyl)-6,7-methylenedioxyquinolin-4-yl-phosphate (I-2-b)

A suspension of compound I-2-a (97.7 mg, 0.18 mmol) in anhydrous MeOH(10 ml) was submitted to hydrogenation in the presence of 10% Pd/C (50mg) at room temperature for 10 min. The catalyst and precipitates wascollected and dissolved in 10% NaHCO₃ solution then filtered. Thefiltrate was acidified with dil HCl, the solid was then collected byfiltration and washed with acetone to give compound I-2-b (60.8 mg,93.1%).

MP>300° C.

¹H-NMR (DMSO-d6, 200 MHz): δ 7.91 (s, 1H, H-2′), 7.87 (s, 1H, H-4′),7.83 (s, 1H, H-5′), 7.50-7.62 (m, 2H, H-5, H-8), 7.25-7.36 (m, 2H, H-5′,H-6′), 6.24 (s, 2H, OCH₂O).

MS (m/z) 362 (ES−)

Anal. calcd for C₁₆H₁₃FNO₆P: C, 52.91; H, 3.05; N, 3.86. Found: C,52.86; H, 3.12; N, 3.79.

Sodium 2-(3′-fluorophenyl)-6,7-methylenedioxyquinoline 4-yl-phosphate(I-2-c)

Compound I-2-b was added to a mixture of 20 ml Amberlite IR-120 (Na⁺form) and 20 ml water, and then stirred for 6 h at room temperature. Themixture was then filtered to remove Amberlite, and then lyophilized togive I-2-c (68.2 mg, 71%).

¹H-NMR (D2O, 200 MHz): δ 7.26-7.78 (m, 5H, H-5, H-8, H-2′, H-5′, H-6′),6.90-6.96 (m, 2H, H-3, H-4′), 6.03 (s, 2H, OCH₂O).

Example 3

Dibenzyl 6-fluoro-2-phenylquinolin-4-yl-phosphate (I-3-a)

Sodium hydride (13.7 mg, 0.57 mmol) was added at 0° C. to a stirredsolution of compound I-3 (55.0 mg, 0.23 mmol) in dry tetrahydrofuran (10ml). After 1 h, tetrabenzyl pyrophosphate (100 mg, 0.19 mmol) was addedand the stirring was continued for 20 min.

The mixture was filtered, and the filtrate was concentrated under vacuumat a temperature below 35° C. The residue was dissolved indichloromethane, washed with an aqueous solution of sodium hydrogencarbonate, dried over MgSO₄ and concentrated under vacuum to give I-3-aas a colorless oil compound (84.4 mg, 89%).

¹H-NMR (DMSO-d6, 200 MHz): δ 8.07-8.14 (m, 1H, H-8), 7.92-7.97 (m, 2H,H-2′, H-6′), 7.67-7.77 (m, 2H, H-3′, H-5′), 7.40-7.50 (m, 10H, Ph), 5.31(s, 2H, —CH₂ -Ph), 5.27 (s, 2H, —CH₂ -Ph)

MS (m/z) 500 (ES+)

Anal. calcd for C₂₉H₂₃FNO₆P: C, 69.74; H, 4.64; N, 2.80. Found: C,69.75; H, 4.60; N, 2.81.

6-Fluoro-2-phenylquinolin-4-yl-phosphate (I-3-b)

A suspension of compound I-3-a (89.8 mg, 0.18 mmol) in anhydrous MeOH(10 ml) was submitted to hydrogenation in the presence of 10% Pd/C (50mg) at room temperature for 10 min. The catalyst and precipitates wascollected and dissolved in 10% NaHCO₃ solution then filtered. Thefiltrate was acidified with dil HCl, the solid was then collected byfiltration and washed with acetone to give compound I-3-b. (50.5 mg,88%).

MP>300° C.

¹H-NMR (DMSO-d6, 200 MHz): δ 8.07-8.14 (m, 3H, H-8, H-2′, H-6′), 7.95(s, 1H, H-5), 7.70-7.74 (m, 2H, H-3′, H-5′), 7.50-7.56 (m, 3H, H-3, H-7,H-4′)

MS (m/z) 318 (ES−)

Anal. calcd for C₁₅H₁₁FNO₄P: C, 56.44; H, 3.47; N, 4.39. Found: C,56.42;

H, 3.49; N, 4.30.

Sodium 6-Fluoro-2-phenylquinolin-4-yl-phosphate (I-3-c)

Compound I-3-b was added to a mixture of 20 ml Amberlite IR-120 (Na⁺form) and 20 ml water, and then stirred for 6 h at room temperature. Themixture was then filtered to remove Amberlite, and then lyophilized togive I-3-c (41.9 mg, 73%).

¹H-NMR (D2O, 200 MHz): δ 7.20-7.83 (m, 5H, H-5, H-7, H-8, H-2′, H-6′),7.25-7.31 (m, 4H, H-3, H-3′, H-4′, H-5′).

Example 4

Dibenzyl 2-phenylquinolin-4-yl-phosphate (I-4-a)

Sodium hydride (13.7 mg, 0.57 mmol) was added at 0° C. to a stirredsolution of compound I-4 (50.8 mg, 0.23 mmol) in dry tetrahydrofuran (10ml). After 1 h, tetrabenzyl pyrophosphate (100 mg, 0.19 mmol) was addedand the stirring was continued for 20 min.

The mixture was filtered, and the filtrate was concentrated under vacuumat a temperature below 35° C. The residue was dissolved indichloromethane, washed with an aqueous solution of sodium hydrogencarbonate, dried over MgSO₄ and concentrated under vacuum to give I-4-aas a colorless oil compound (71.3 mg, 78%).

¹H-NMR (DMSO-d6, 200 MHz): δ 8.05 (d, J=8.2 Hz, 1H, H-5), 7.73-7.98 (m,5H, H-6, H-7, H-8, H-2′, H-6′), 7.58 (d, J=8.0 Hz, 1H, H-4′), 7.48-7.51(m, 3H, H-3, H-3′, H-5′), 7.29-7.40 (m, 10H, Ph), 5.31 (s, 2H, —CH₂-Ph), 5.27 (s, 2H, —CH₂ -Ph)

MS (m/z) 482 (ES+)

Anal. calcd for C₂₉H₂₄NO₆P: C, 72.34; H, 5.02; N, 2.90. Found: C, 71.89;H, 5.13; N, 2.88.

2-Phenylquinolin-4-yl-phosphate (I-4-b)

A suspension of compound I-4-a (86.6 mg, 0.18 mmol) in anhydrous MeOH(10 ml) was submitted to hydrogenation in the presence of 10% Pd/C (50mg) at room temperature for 10 min. The catalyst and precipitates wascollected and dissolved in 10% NaHCO₃ solution then filtered. Thefiltrate was acidified with dil HCl, the solid was then collected byfiltration and washed with acetone to give compound I-4-b (48.9 mg,90.3%).

MP>300° C.

¹H-NMR (DMSO-d6, 200 MHz): δ 7.80-8.12 (m, 4H, H-5, H-8, H-2′, H-6′),7.49-7.78 (m, 6H, H-3, H-6, H-7, H-3′, H-4′, H-5′), 7.78 (s, 1H, H-7),7.66 (t, J=8.0 Hz), 7.42-7.50 (m, 4H, H-3, H-3′, H-4′, H-5′)

MS (m/z) 300 (ES−)

Anal. calcd for C₁₅H₁₂NO₆P: C, 59.81; H, 4.02; N, 4.65. Found: C, 59.52;H, 4.13; N, 4.72.

Sodium 6-fluoro-2-phenylquinolin-4-yl-phosphate (I-4-c)

Compound I-4-b was added to a mixture of 20 ml Amberlite

IR-120 (Na⁺ form) and 20 ml water, and then stirred for 6 h at roomtemperature. The mixture was then filtered to remove Amberlite, and thenlyophilized to give I-4-c (41.2 mg, 74%).

¹H-NMR (D2O, 200 MHz): δ 8.21 (d, J=8.2 Hz, 1H, H-5), 7.80-7.89 (m, 3H,H-8, H-2′, H-6′), 7.78 (s, 1H, H-7), 7.66 (t, J=8.0 Hz), 7.42-7.50 (m,4H, H-3, H-3′, H-4′, H-5′)

Example 5

Dibenzyl 6-methoxy-2(3-'fluorophenyl)-quinolin-4-yl-phosphate (I-5-a)

Sodium hydride (13.7 mg, 0.57 mmol) was added at 0° C. to a stirredsolution of compound I-5 (61.9 mg, 0.23 mmol) in dry tetrahydrofuran (10ml). After 1 h, tetrabenzyl pyrophosphate (100 mg, 0.19 mmol) was addedand the stirring was continued for 20 min.

The mixture was filtered, and the filtrate was concentrated under vacuumat a temperature below 35° C. The residue was dissolved indichloromethane, washed with an aqueous solution of sodium hydrogencarbonate, dried over MgSO₄ and concentrated under vacuum to give I-5-aas a colorless oil compound (85.4 mg, 85%)

¹H-NMR (DMSO-d6, 200 MHz): δ 7.98 (d, J=9.4 Hz, 1H, H-8), 7.74-7.83 (m,3H, H-5, H-7, H-5′), 7.43-7.54 (m, 1H, H-6′), 7.41-7.48 (m, 1H, H-2′),7.20-7.22 (m, H-3), 5.31 (s, 2H, —CH₂ -Ph), 5.27 (s, 2H, —CH₂ -Ph), 3.78(s, 3H, OCH₃).

MS (m/z) 530 (ES+)

Anal. calcd for C₃₀H₂₅FNO₅P: C, 68.05; H, 4.76; N, 2.65. Found: C,67.32; H, 4.33; N, 2.78.

6-Methoxy-2(3-′fluorophenyl)-quinolin-4-yl-phosphate (I-5-b)

A suspension of compound I-5-a (95.2 mg, 0.18 mmol) in anhydrous MeOH(10 ml) was submitted to hydrogenation in the presence of 10% Pd/C (50mg) at room temperature for 10 min. The catalyst and precipitates wascollected and dissolved in 10% NaHCO₃ solution then filtered. Thefiltrate was acidified with dil HCl, the solid was then collected byfiltration and washed with acetone to give compound I-5-b (56.5 mg,89.9%).

MP>300° C.

¹H-NMR (DMSO-d6, 200 MHz): δ 7.93-7.89 (m, 4H, H-5, H-7, H-8, H-5′),7.45-7.58 (m, 1H, H-6′), 7.35-7.41 (m, 2H, H-2′, H-4′), 7.20-7.32 (m,1H, H-3), 3.81 (s, 3H, OCH₃)

MS (m/z) 348 (ES−)

Anal. calcd for C₁₆H₁₃FNO₅P: C, 55.02; H, 3.75; N, 4.01. Found: C,54.90; H, 3.89; N, 4.35.

Example 6

Dibenzyl 2-(3′-methoxyphenyl)-6,7-methylenedioxyquinolin-4-yl-phosphate(I-6-a)

Sodium hydride (13.7 mg, 0.57 mmol) was added at 0° C. to a stirredsolution of compound I-6 (67.9 mg, 0.23 mmol) in dry tetrahydrofuran (10ml). After 1 h, tetrabenzyl pyrophosphate (100 mg, 0.19 mmol) was addedand the stirring was continued for 20 min.

The mixture was filtered, and the filtrate was concentrated under vacuumat a temperature below 35° C. The residue was dissolved indichloromethane, washed with an aqueous solution of sodium hydrogencarbonate, dried over MgSO₄ and concentrated under vacuum to give togive I-6-a as a colorless oil compound (88.6 mg, 84%)

¹H-NMR (DMSO-d6, 200 MHz): δ 7.60 (s, 1H, H-6′), 7.55 (s, 1H, H-2′),7.25-7.40 (m, 14H, I-1-5, H-8, H-4′, H-5′, Ph), 6.21 (s, 2H, OCH₂O),5.28 (s, 2H, —CH₂ -Ph), 5.24 (s, 2H, —CH₂ -Ph), 3.80 (s, 3H, OCH₃)

MS (m/z) 556 (ES+)

Anal. calcd for C₃₁H₂₆NO₇P: C, 67.02; H, 4.72; N, 2.52. Found: C, 68.15;H, 4.68; N, 2.61.

2-(3′-Methoxyphenyl)-6,7-methylenedioxyquinolin-4-yl-phosphate (I-6-b)

A suspension of compound I-6-a (97.74 mg, 0.18 mmol) in anhydrous MeOH(10 ml) was submitted to hydrogenation in the presence of 10% Pd/C (50mg) at room temperature for 10 min. The catalyst and precipitates wascollected and dissolved in 10% NaHCO₃ solution then filtered. Thefiltrate was acidified with dil HCl, the solid was then collected byfiltration and washed with acetone to give compound I-6-b (63.5 mg,94%).

MP>300° C.

MS (m/z) 374 (ES−)

Anal. calcd for C₁₇H₁₄NO₇P: C, 54.41; H, 3.76; N, 3.73. Found: C, 53.86;H, 3.66; N, 3.81.

Sodium 2-(3′-methoxyphenyl)-6,7-methylenedioxyquinolin-4-yl-phosphate(I-6-c)

Compound I-6-b was added to a mixture of 20 ml Amberlite IR-120 (Na⁺form) and 20 ml water, and then stirred for 6 h at room temperature. Themixture was then filtered to remove Amberlite, and then lyophilized togive I-6-c (53.9 mg, 76%).

¹H-NMR (D2O, 200 MHz): δ 7.56 (s, 1H, H-6′), 7.25-7.42 (m, 4H, H-5, H-8,H-2′, H-5′), 7.12 (s, 1H, H-4′), 6.95 (s, 1H, H-3), 6.00 (s, 2H, OCH₂O),3.62 (s, 3H, OCH₃)

In the following Example 7, a novel intermediate, 2-selenophene4-quinolone (I-7-d), was synthesized. 2-selenophene-4-quinolone (I-7-d)was reacted with tetrabenzyl pyrophosphate in the presence of alkali,the corresponding phosphoric acid dibenzyl ester (I-7-e) was obtained.

Example 7

Selenophene-2-carboxylic acid (I-7-a)

To a solution of selenophene (20 g, 152.7 mmol) in (Et)₂O (150 ml) wasadded TMEDA (25.5 ml, 170.0 mmol) and n-butyllithium (66.1 ml of a 2.5 Msolution in hexane, 152.8 mmol). The resulting solution was heated atreflux for 1.5 h, and then cooled in an acetone/CO₂ bath, after whichcrushed solid carbon dioxide (40 g, 909.1 mmol) was added. The reactionmixture was allowed to return to room temperature, and quenched byaddition of 10% KOH solution. The aqueous layer was acidified to pH 3with 8 M HCl, extracted with (Et)₂O, washed with brine, dried over MgSO₄filtered and concentrated under vacuum to give compound I-7-a (24.6 g,92.1%).

MP 122-124° C.

¹H-NMR (CDCl₃-d₁, 200 MHz): δ 8.92 (s, 1H, —COOH), 8.37 (dd, J=1.0 Hz,5.6 Hz, 1H, H-3), 8.13 (dd, J=0.8 Hz, 3.8 Hz, 1H, H-5), 7.37 (dd, J=3.8Hz, 5.6 Hz, 1H, H-4).

MS (m/z) 175.0 (EI+)

Anal. calcd for C₅H₄O₂Se: C, 34.31; H, 2.30. Found: C, 34.33; H, 2.28.

N-(5-acetylbenzo[d][1,3]dioxol-6-yl)selenophene-2-carboxamide (I-7-c)

I-7-a (2 g, 11.40 mmol) was taken for subsequent chlorination byrefluxing with thionyl chloride (4.1 ml, 56.18 mmol) for 20 h to affordI-7-b, which, without further purification, was treated with2-amino-(4,5-methylenedioxy)-acetophenone (1.63 g, 9.12 mmol) andtriethylamine (2 ml, 14.80 mmol) in 100 ml toluene, and refluxed for 3h. The reaction mixture was concentrated under vacuum, and the solidmaterial is consecutively washed with ethanol and dried at 80° C. for 2h to give crude compound I-7-c (2.7 g, 74%).

MP 198.5-198.8° C.

¹H-NMR (DMSO-d6, 200 MHz): δ 12.85 (s, 1H, NHCO), 8.52 (d, J=5.1 Hz, 1H,H-3′), 8.16 (s, 1H, H-4), 7.93 (d, J=3.8 Hz, 1H, H-5′), 7.61 (s, 1H,H-7), 7.49-7.46 (m, 1H, H-4′), 6.13 (s, 2H, OCH₂O), 2.58 (s, 3H, CH₃).

MS (m/z) 336.2 (EI+)

Anal. calcd for C14H11NO4Se: C, 50.01; H, 3.30; N, 4.17. Found: C,50.11; H, 3.32; N, 4.15.

2-(2′-Selenophenyl)-6,7-(methylenedioxy)-4-quinolone (I-7-d)

I-7-c (2.7 g, 8.0 mmol) was suspended in 100 ml t-BuOH. Potassiumtert-butoxide (4.49 g, 40 mmol) was added, and the mixture was heated atreflux for 24 h. The mixture was cooled to room temperature, and pouredonto 100 ml of aqueous NH₄Cl. The yellow-brown solid was collected andwashed by distilled water to give compound I-7-d (3.1 g, 85%).

MP>300° C.

¹H-NMR (DMSO-d6, 200 MHz): δ 8.27 (s, 1H, H-3′), 7.83 (s, 1H, H-5′),7.39 (t, J=4.5 Hz, 1H, H-4′), 7.31 (s, 1H, H-5), 7.14 (s, 1H, H-8), 6.11(s, 3H, H-3, OCH₂O).

MS (m/z) 318.2 (EI+)

Anal. calcd for C14H9NO3Se: C, 52.85; H, 2.85; N, 4.40. Found: C, 52.87;H, 2.82; N, 4.45.

Dibenzyl 2-(2′-selenophenyl)-6,7-methylenedioxyquinolin-4-yl-phosphate(I-7-e)

Sodium hydride (30 mg, 1.25 mmol) was added at 0° C. to a stirredsolution of compound I-7-d (100.0 mg, 0.32 mmol) in dry tetrahydrofuran(10 ml). After 1 h, tetrabenzyl pyrophosphate (204.6 mg, 0.38 mmol) wasadded and the stirring was continued for 20 min.

The mixture was filtered, and the filtrate was concentrated under vacuumat a temperature below 35° C. The residue was dissolved indichloromethane, washed with an aqueous solution of sodium hydrogencarbonate, dried over MgSO₄ and concentrated under vacuum to give thesolid which was subjected to silica gel column chromatography. Elutionwith CH₂Cl₂ gave yellowish compound I-7-e (151.8 mg, 82%).

MP 110.5-110.8° C.

¹H-NMR (DMSO-d6, 200 MHz): δ 8.24 (d, J=5.6 Hz, 1H, H-3′), 7.65 (d,J=3.8 Hz, 1H, H-5′), 7.57 (s, 1H, H-5), 7.05 (s, 1H, H-8), 7.39-7.26 (m,11H, H-4′, Ph), 6.19 (s, 2H, OCH₂O), 5.28 (s, 2H, —CH₂ -Ph), 5.24 (s,2H, —CH₂ -Ph).

MS (m/z) 580 (ES+)

Anal. calcd for 28₀H₂₂NO₆PSe: C, 58.14; H, 3.83; N, 2.42. Found: C,57.28; H, 3.56; N, 2.59.

Example 8

6-Methyl-2-phenylquinolin-4(1H)-one (I-8-a)

A mixture of p-toluidine (2.14 g, 0.02 mole), ethyl benzoylacetate (4.9g, 0.025 mole), and polyphosphoric acid (PPA) was heated at 130° C. withstirring. After the reaction was complete, the mixture was cooled toroom temperature and neutralized with 4 M NaOH. The yellow solid wasfiltered, washed with water, dried and recrystallized from ethanol togive compound I-8-a as white solid (2.9 g, 48.9%).

MP 290.2-291.5° C.

¹H-NMR (DMSO-d6, 200 MHz): δ 11.55 (1H, s, H-1), 7.88 (1H, s, H-5),7.79-7.82 (2H, m, H-2′, H-3′), 7.66 (1H, d, J=8.5 Hz, H-8), 7.54-7.57(3H, m, H-3′, H-4′, H-5′), 7.48 (1H, d, J=8.5 Hz, H-7), 6.31 (1H, s,H-3), 2.40 (3H, s, CH₃)

MS (m/z) 235 (EI+)

Anal. calcd for C16H13NO: C, 81.68; H, 5.57; N, 5.95. Found: C, 81.60;H, 5.63; N, 5.88.

4-(Benzyloxy)-6-methyl-2-phenylquinoline (I-8-b)

I-8-a (700 mg, 3 mmole) was dissolved in dry DMF (30 ml), and NaH (360mg, 15 mmole) was added protionwise with stirring for 30 min at roomtemperature. Benzyl chloride (750 mg, 6 mmole) was then added dropwise,and stirred at room temperature overnight. The reaction mixture waspoured into ice-water and extracted with CH₂Cl₂. The organic layer waswashed with water, dried over MgSO₄, and evaporated. The residue wasfurther chromatographed over silica gel by elution with n-hexane-EtOAc(3:1), and recrystallized from n-hexane-CH₂Cl₂ to afford I-8-b as whitecrystal (536 mg, 54.9%).

MP 138.6-139.3° C.

¹H-NMR (DMSO-d6, 200 MHz): δ 8.23-8.26 (2H, m, H-2′, H-6′), 7.88-7.91(2H, m, H-5, H-8), 7.37-7.62 (9H, m, H-7, H-3′, H-4′, H-5′, Ph), 5.51(2H, s, OCH ² Ph), 2.48 (3H, s, CH₃)

Anal. calcd for C23H19NO: C, 84.89; H, 5.89; N, 4.30. Found: C, 84.93;H, 5.85; N, 4.33.

N-{[4-(Benzyloxy)-2-phenylquinolin-6-yl]methyl}-N-ethyl ethanamine(I-8-d)

I-8-b (650 mg, 2 mmol), N-bromo-succinimide (NBS, 360 mg, 2 mmol), and2,2′-azobis(isobutyronitrile) (AIBN, 30 mg, 0.19 mmol) were added to adry round bottom flask, which was purged with argon. 50 ml of drybenzene was added to the reaction mixture in an argon atmosphere withstirring at room temperature for 30 min, and then refluxed at 80° C. for1 h and then cooled to room temperature to give I-7-c, which, withoutfurther purification, was treated with diethylamine (3.0 ml, 29.0mmole), and then refluxed for 1 h. After removing the solvent byevaporation, the mixture was partitioned with EtOAc and 50 ml 10% HCl.,and then the acid layer was neutralized to PH 7-8 by 10% NaHCO₃,extracted with EtOAc (100 ml×5). The organic layer was dried over MgSO₄,and evaporated. The residue was further chromatographed over silica gelby elution with CH₂Cl₂-methanol (3:1), and recrystallized fromn-hexane-EtOAc to afford I-8-d as light-yellow solid (120 mg, 15.1%).

MP 107.7-108.6° C.

¹H-NMR (DMSO-d6, 200 MHz): δ 8.22 (2H, m, H-2′, H-6′), 8.01 (1H, s,H-5), 7.91 (1H, d, H-8), 7.33-7.69 (9H, m, H-7, H-3′, H-4′, H-5′, Ph),5.49 (2H, s, OCH₂Ph), 3.65 (2H, s, CH₂N(CH₂CH₃)₂), 2.43 (4H, q, J=7 Hz,CH₂N(CH ² CH₃)₂), 0.93 (6H, t, J=7 Hz, CH₂N(CH₂CH₃) ² )

MS (m/z) 396 (EI+)

Anal. calcd for C27H28N2O: C, 81.78; H, 7.12; N, 7.06. Found: C, 81.68;H, 7.03; N, 7.15.

6-[(Diethylamino)methyl]-2-phenylquinolin-4(1H)-one (I-8-e)

I-8-d (120 mg, 0.3 mmol) was dissolved in glacial acetic acid (5 ml).HBr (3 ml) was added while the solution was heated to 60° C., and themixture was heated to 90° C. for 3 h. After the reaction was complete,the reaction mixture was poured into water, and extracted with EtOAc.The acid layer was neutralized to pH 7-8 by adding 10% NaHCO₃, andextracted with EtOAc (100 ml×5). The organic layer was dried over MgSO₄,and evaporated. The residue was recrystallized from n-hexane-EtOAc toafford I-8-d as gray solid (55 mg, 59.9%).

MP 227.9-229.7° C.

¹H-NMR (DMSO-d6, 200 MHz): δ 7.96 (1H, s, H-5), 7.78 (2H, in, H-2′,H-6′), 7.69 (1H, d, H-8), 7.50-7.58 (4H, m, H-7, H-3′, H-4′, H-5′), 6.31(1H, s, H-3), 3.55 (2H, s, CH ² N(CH₂CH₃)₂), 2.41 (4H, q, J=7 Hz,CH₂N(CH ² CH₃)₂), 0.92 (6H, t, J=7 Hz, CH₂N(CH₂CH₃) ² )

MS (m/z) 306 (EI+)

Anal. calcd for C20H22N2O: C, 78.40; H, 7.24; N, 9.14. Found: C, 78.43;H, 7.35; N, 9.08.

Example 9

2-(2-Fluorophenyl)-6-methylquinolin-4(1H)-one (I-9-a)

A mixture of p-toluidine (2.14 g, 0.02 mole), 2-fluoro-ethylbenzoylacetate (5.25 g, 0.025 mole), and polyphosphoric acid (PPA) washeated at 130° C. with stirring. After the reaction was complete, themixture was cooled to room temperature and neutralized with 4 M NaOH.The yellow solid was filtered, washed with water, dried andrecrystallized from ethanol to give compound I-9-a as white solid (2.6g, 51.3%).

MP 259.1-259.9° C.

¹H-NMR (DMSO-d6, 200 MHz): δ7.86 (1H, s, H-5), 7.64 (1H, td, J=7.58,H-4′), 7.47-7.57 (3H, m, H-7, H-8, H-6′), 7.30-7.43 (2H, d, J=7.02, dd,J=7.36, H-3′, 5′), 6.12 (1H, s, H-3), 2.36 (3H, s, CH₃)

MS (m/z) 253 (EI+)

Anal. calcd for C16H22FNO: C, 75.88; H, 4.78; N, 5.53. Found: C, 75.94;H, 4.70; N, 5.46.

4-(Benzyloxy)-2-(2-fluorophenyl)-6-methylquinoline (I-9-b)

I-9-a (750 mg, 3 mmole) was dissolved in dry DMF (30 ml), and NaH (360mg, 15 mmole) was added protionwise with stirring for 30 min at roomtemperature. Benzyl chloride (750 mg, 6 mmole) was then added dropwise,and stirred at room temperature overnight. The reaction mixture waspoured into ice-water and extracted with CH₂Cl₂. The organic layer waswashed with water, dried over MgSO₄, and evaporated. The residue wasfurther chromatographed over silica gel by elution with n-hexane-EtOAc(3:1), and recrystallized from n-hexane-CH₂Cl₂ to afford I-9-b as whitecrystal (515 mg, 50.0%).

MP 91.5-92.8° C.

¹H-NMR (DMSO-d6, 200 MHz): δ 7.84-7.97 (3H, m, H-5, H-8, H-4′),7.26-7.58 (10H, m, H-3, H-7, H-3′, H-5′, H-6′, Ph), 5.38 (2H, s, OCH ²Ph), 2.45 (3H, s, CH₃)

MS (m/z) 343 (EI+)

Anal. calcd for C23H18FNO: C, 80.45; H, 5.28; N, 4.08. Found: C, 80.51;H, 5.29; N, 4.17.

N-{[4-(Benzyloxy)-2-(2-fluorophenyl)quinolin-6-yl]methyl}-N-ethylethanamine(I-9-d)

I-9-b (680 mg, 2 mmol), N-bromo-succinimide (NBS, 360 mg, 2 mmol), and2,2′-azobis(isobutyronitrile) (AlBN, 30 mg, 0.19 mmol) were added to adry round bottom flask, which was purged with argon. 50 ml of drybenzene was added to the reaction mixture in an argon atmosphere withstirring at room temperature for 30 min, and then refluxed at 80° C. for1 h and then cooled to room temperature to give I-9-c, which, withoutfurther purification, was treated with diethylamine (3.0 ml, 29.0mmole), and then refluxed for 1 h. After removing the solvent byevaporation, the mixture was partitioned with EtOAc and 50 ml 10% HCl.,and then the acid layer was neutralized to PH 7-8 by 10% NaHCO₃,extracted with EtOAc (100 ml×5). The organic layer was dried over MgSO₄,and evaporated. The residue was further chromatographed over silica gelby elution with CH₂Cl₂-methanol (3:1), and recrystallized fromn-hexane-EtOAc to afford I-9-d as yellow solid (120 mg, 15.1%).

MP 51.2-51.5° C.

¹H-NMR (DMSO-d6, 200 MHz): δ 8.04 (1H, s, H-5), 7.84-7.96 (2H, m, H-8,H-5′), 7.69 (1H, dd, H-4′), 7.28-7.54 (9H, m, H-3, H-7, H-3′, H-6′, Ph),5.41 (2H, s, OCH ² Ph), 3.68 (2H, s, CH ² N(CH ² CH₃)₂), 2.46 (4H, q,J=7, CH₂N(CH ² CH₃)₂), 0.94 (6H, t, J=7, CH₂N(CH₂CH₃) ² )

MS (m/z) 414 (EI+)

Anal. calcd for C27H27FN2O: C, 78.23; H, 6.57; N, 6.76. Found: C, 78.25;H, 6.67; N, 6.74.

6-[(Diethylamino)methyl]-2-(2-fluorophenyl)quinolin-4(1H)-one (I-9-e)

I-9-d (120 mg, 0.3 mmol) was dissolved in glacial acetic acid (5 ml).HBr (3 ml) was added while the solution was heated to 60° C., and themixture was heated to 90° C. for 3 h. After the reaction was complete,the reaction mixture was poured into water, and extracted with EtOAc.The acid layer was neutralized to pH 7-8 by adding 10% NaHCO₃, andextracted with EtOAc (100 ml×5). The organic layer was dried over MgSO₄,and evaporated. The residue was recrystallized from n-hexane-EtOAc toafford I-8-e as gray solid (58 mg, 59.6%). MP 184.2-184.7° C.

¹H-NMR (DMSO-d6, 200 MHz): δ 11.9 (1H, s, H-1), 7.97 (1H, s, H-5),7.52-7.69 (41-1, m, H-7, H-8, H-4′, H-6′), 7.31-7.43 (2H, m, H-3′,H-5′), 6.12 (1H, s, H-3), 3.57 (2H, s, CH₂N(CH₂CH₃)₂), 2.40 (4H, q, J=7Hz, CH₂N(CH₂CH₃)₂), 0.92 (6H, t, J=7 Hz, CH₂N(CH₂CH₃)₂)

MS (m/z) 324 (EI+)

Anal. calcd for C20H21FN2O: C, 74.05; H, 6.53; N, 8.64. Found: C, 73.94;H, 6.62; N, 8.67.

Example 10

Ethyl 3-methyl-benzoyl-acetate (I-10-a)

To a vigorously stirred suspension of NaH (564 mg, 48.5 mmol) andCO(OEt)₂ (5.73 g, 48.5 mmol) in anhydrous toluene (50 ml) was addeddropwise a solution of 3-methylacetophenone (4.33 g, 32.3 mmole) intoluene under reflux. The mixture was allowed to reflux and was stirredfor 30 min after the addition was complete. When cooled to roomtemperature, the mixture was acidified with glacial AcOH. After ice-coldwater was added, the mixture was extracted with toluene. The organiclayer was dried over MgSO₄, and evaporated. The residue was furtherchromatographed over silica gel by elution with CH₂Cl₂-n-haxane (3:2) toafford I-10-b as light-yellow liquid (3.13 g, 46.9%)

¹H-NMR (DMSO-d6, 200 MHz): δ 7.68-7.72 (2H, m, H-4, H-6), 7.32-7.36 (2H,m, H-2, H-3), 4.16 (2H, q, J=7, CH ² CH₃), 3.94 (2H, s, H-10), 2.38 (3H,s, CH₃), 1.2 (3H, t, J=7, CH₂CH ³ )

MS (m/z) 206 (EI+)

Anal. calcd for C12H14O3: C, 69.88; H, 6.84. Found: C, 69.72; H, 6.95.

6-Methoxy-2-m-tolylquinolin-4(1H)-one (I-10-b)

A mixture of p-anisidine (2.14 g, 0.02 mole), I-10-a (5.1 g, 0.025mole), and polyphosphoric acid (PPA) was heated at 130° C. withstirring. After the reaction was complete, the mixture was cooled toroom temperature and neutralized with 4 M NaOH. The yellow solid wasfiltered, washed with water, dried and recrystallized from ethanol togive compound I-9-a as light-purple solid (2.6 g, 25.8%).

MP 262.2-264.1° C.

¹H-NMR (DMSO-d6, 200 MHz): δ 7.70 (1H, d, H-8), 7.55-7.60 (2H, m, H-5,7), 7.25-7.47 (4H, m, H-2′, H-4′, H-5′, H-6′), 6.33 (1H, s, H-3), 3.80(3H, s, OCH₃), 2.37 (3H, s, CH₃)

MS (m/z) 265 (EI+)

Anal. calcd for C17H15NO: C, 76.79; H, 5.70; N, 5.28. Found: C, 76.81;5.62; N, 5.34.

4-(Benzyloxy)-6-methoxy-2-m-tolylquinoline (I-10-c)

I-10-b (795 mg, 3 mmole) was dissolved in dry DMF (30 ml), and NaH (360mg, 15 mmole) was added protionwise with stirring for 30 min at roomtemperature. Benzyl chloride (750 mg, 6 mmole) was then added dropwise,and stirred at room temperature overnight. The reaction mixture waspoured into ice-water and extracted with CH₂Cl₂. The organic layer waswashed with water, dried over MgSO₄, and evaporated. The residue wasfurther chromatographed over silica gel by elution with n-hexane-EtOAc(3:1), and recrystallized from n-hexane-CH₂Cl₂ to afford I-10-c as whitecrystal (530 mg, 49.7%).

MP 133.0-134° C.

¹H-NMR (DMSO-d6, 200 MHz): δ 8.00 (1H, s, H-5), 7.96 (1H, d, H-8), 7.89(1H, d, J=8 Hz, H-7), 7.32-7.58 (6H, m, H-3, H-2′, H-5′, H-6′, Ph), 7.22(1H, d, J=7 Hz, H-4′), 5.50 (2H, s, OCH ² Ph), 3.83 (3H, s, OCH3), δ2.38(3H, s, CH₃)

MS (m/z) 355 (EI+)

Anal. calcd for C24H21NO2: C, 81.10; H, 5.96; N, 3.94. Found: C, 81.9;H, 5.81; N, 3.97.

N-{[3-(4-(Benzyloxy)-6-methoxyquinolin-2-yl)phenyl)methyl}-N-ethylethanamine(I-10-e)

I-10-c (530 mg, 2 mmol), N-bromo-succinimide (NBS, 360 mg, 2 mmol), and2,2′-azobis(isobutyronitrile) (AIBN, 30 mg, 0.19 mmol) were added to adry round bottom flask, which was purged with argon. 50 ml of drybenzene was added to the reaction mixture in an argon atmosphere withstirring at room temperature for 30 min, and then refluxed at 80° C. for1 h and then cooled to room temperature to give I-10-d, which, withoutfurther purification, was treated with diethylamine (3.0 ml, 29.0mmole), and then refluxed for 1 h. After removing the solvent byevaporation, the mixture was partitioned with EtOAc and 50 ml 10% HCl.,and then the acid layer was neutralized to PH 7-8 by 10% NaHCO₃,extracted with EtOAc (100 ml×5). The organic layer was dried over MgSO₄,and evaporated. The residue was further chromatographed over silica gelby elution with CH₂Cl₂-methanol (3:1), and recrystallized fromn-hexane-EtOAc to afford I-10-e as yellow solid (25 mg, 2.9%).

MP 89.2-89.5° C.

¹H-NMR (DMSO-d6, 200 MHz): δ 8.13 (1H, s, H-3). 7.87-8.04 (2H, m, H-7,8), 7.34-7.43 (10H, m, H-3, H-2′, H-4′, H-5′, H-6′, Ph), 5.51 (2H, s,OCH ² Ph), 3.84 (3H, s, OCH₃), 3.69 (2H, s, CH ² N(CH₂CH₃)₂), 2.53 (4H,q, J=7 Hz, CH₂N(CH ² CH₃)₂), 0.99 (6H, t, J=7 Hz, CH₂N(CH₂CH₃) ² )

MS (m/z) 426 (EI+)

Anal. calcd for C28H30N2O2: C, 78.83; H, 7.90; N, 6.57. Found: C, 78.95;H, 7.14; N, 6.48.

2-{3-[(Diethylamino)methyl]phenyl}-6-methoxyquinolin-4(1H)-one (I-10-f)

I-10-e (42 mg, 0.1 mmol) was dissolved in glacial acetic acid (5 ml).HBr (3 ml) was added while the solution was heated to 60° C., and themixture was heated to 90° C. for 3 h. After the reaction was complete,the reaction mixture was poured into water, and extracted with EtOAc.The acid layer was neutralized to pH 7-8 by adding 10% NaHCO₃, andextracted with EtOAc (100 ml×5). The organic layer was dried over MgSO₄,and evaporated. The residue was recrystallized from n-hexane-EtOAc toafford I-10-f as gray solid (20.8 mg, 61.9%).

MP 152.1-152.7° C.

¹H-NMR (DMSO-d6, 200 MHz): δ 11.76 (1H, s, H-1), 7.67-7.74 (3H, m, H-5,H-8, H-6′), 7.46-7.49 (3H, m, H-7, H-2′, H-4′), 7.27 (1H, dd, H-5′),6.27 (1H, s, H-3), 3.80 (3H, s, OCH₃), 3.67 (2H, s, CH ² N(CH₂CH₃)₂),2.53 (4H, q, J=7 Hz, CH₂N(CH ² CH₃)₂), 0.97 (6H, t, J=7 Hz, CH₂N(CH₂CH₃)² ).

Anal. calcd for C21H24N2O2: C, 74.97; H, 7.19; N, 8.33. Found: C, 74.81;H, 7.33; N, 8.31.

Anti Cancer Activities Effects of Compounds I-1 and I-1-b on Anti-TumorActivity In Vivo (I) Effects of Compounds I-1 and I-1-b on MCF-7 TumorXenograft Model I-1 Materials and Methods

Female GALB/cAnN-Foxn1.E SCID mice (18-20 g; 6-8 weeks of age) werepurchased from the National Animal Center and maintained in pressurizedventilated cage according to institutional regulations. The mice wereimplanted subcutaneously with estradiol (0.7 mg) 2 days before tumortransplantation. MCF-7 cells (2×10⁶) were inoculated s.c. into the rightflank of the mice. After appearance of a 150-mm³ tumor nodule, 30tumor-bearing mice were randomly divided into five groups for treatmentwith vehicle (PBS), I-1 or I-1-b. The first groups only receivedvehicle. The second to fifth groups were given i.p. the followingtreatments three times per week, respectively: I-1 (15 mg/kg), I-1 (30mg/kg), I-1-b (22.5 mg/kg), and I-1-b (45 mg/kg). Mice were weighed andtumors were measured using calipers every week. Tumor size wascalculated with the following formula: (L+W)/2, where L is the lengthand W is the width. On the final day of the treatment, mice weresacrificed; tumors were excised, weighted, and sectioned; and the tumorsections were embedded in OCT compound and frozen at −70° C.

I-2 Results

The effects of I-1 or I-1-b, were examined in vivo. Thirty female SCIDmice were individually injected s.c. with MCF7 cells. The mice weredivided into five groups (six mice per group) and treated with vehiclealone, I-1 (15 or 30 mg/kg), I-1-b (22.5 or 45 mg/kg). As shown in FIG.1, this in vivo tumor model shows a significant reduction in tumorvolume in mice treated with 45 mg/kg I-1-b when compared with controlmice (P<0.001). These results demonstrate that I-1-b significantlyinhibited MCF7 tumor growth in a mouse xenograft model.

(II) Effects of Compounds I-1 and I-1-b on CT-26 Intraperitoneal TumorModel II-1 Materials and Methods

30 male 6-week-old Balb/c mice, were purchased from the National Animalcancer and maintained in pressurized ventilated cage according toinstitutional regulations. CT-26 (1×10⁶) cells were injected intoperitoneal cavities at day 0. Animals were randomly assigned toanti-tumoral efficacy study (n=10). Seven days after tumor inoculation,oral administration of 5 and 10 mg/kg of I-1-b (QD for seven times) tothe mice was carried out. The survival rate and body weight of theanimals was monitored.

II-2 Results

II-2-1 Appearance of Mice after Treatment

Mice in the excipient control group showed overt ascites, while micereceiving orally I-1-b (5 mg/kg/day, QD×7) and I-1-b (10 mg/kg/day,QD×7) exhibited reduced ascites development.

II-2-2 the Average Life Span of Mice after Treatment

As shown in FIG. 2, all mice in the excipient control group were dead 40days after, while those receiving compounds I-1-b (5 mg/kg/day, QD×7)and compounds I-1-b (10 mg/kg/day, QD×7) were all dead respectively byday 45 and day 50 post challenge. The average life span was prolonged by140% at the dose of (10 mg/kg/day, QD×7) and by 120% at the dose of (5mg/kg/day, QD×7). A maximally tolerated dose was not achieved.

Cell Viability Assay (MTT Assay)

Cells were seeded in a 24-well microtiter plate (2×10⁴ cells/well)overnight, then treated with DMSO (Control) or various concentrations oftest compounds, and incubated for 48 hours. The effect of test compoundson cell growth was examined by the MTT(3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) assay.Briefly, 40 μl of MTT solution (2 mg/ml, Sigma Chemical Co.) was addedto each well to make a final volume of 500 μl and incubated for 1 h at37° C. The supernatant was aspirated, and the MTT-formazan crystalsformed by metabolically viable cells were dissolved in 200 μl of DMSO.Finally, the absorbance at O.D. 550 nm was detected by enzyme-linkedimmunosorbent assay (ELISA) reader.

Results: Cytotoxic Effect of Compounds I-1-b, I-2-b, I-3-b, I-4-b,I-5-b, I-7-d, I-7-e Against the Human Breast Cancer MCF-7 Cells

The cytotoxic effect of compounds I-1-b, I-2-b, I-3-b, I-4-b, I-5-b,I-7-d, I-7-e were evaluated in the human breast cancer MCF-7 cells. Asshown in FIG. 3, treatment with 0.125 to 10 μM of these compounds causeda dose-dependent decrease of cell viability. These results indicate thatcompounds I-1-b, I-2-b, I-3-b, I-4-b, I-5-b, I-7-d, I-7-e showsignificant cytotoxicity against MCF-7 cells. Therefore, these newderivatives of 2-aryl-quinolines are proposed as potential therapeuticagents for the treatment of cancers.

Cytotoxic Activity of Compound I-7-d

In vitro cytotoxic activity of compound I-7-d was tested in HCT-116, HepG2, NCI-H226, A549, A498 and HL-60 cells. As shown in Table 1, compoundI-7-d demonstrates significant inhibition against most of the six cancercell lines and most notably, is quite active against HCT-116 and HL-60cells. Compound I-7-d shows an IC₅₀ of 0.9 μM against HCT-116 and anIC₅₀ of 0.5 μM against HL-60 cell. Compound I-7-d is an attractivecandidate for development as a novel anti-cancer agent.

TABLE 1 IC₅₀ (μM) HCT116 Hep G2 NCI-H226 A549 A498 HL-60 I-7-d 0.9 4.14.9 8.1 2.7 0.5 * Six cancer cell lines were treated with compound I-7-dfor 48 h. After treatment, cells were harvested and examined using MTTassay. * IC₅₀ value means the concentration causing 50%growth-inhibitory effect. * HCT-116, colon cancer cell line; Hep G2,hepatoma cancer cell line; NCI-H226, non-small cell lung cancer cellline; A549, lung cancer cell line; A498, renal cancer cell line; HL-60,leukemia cancer cell line.

Cytotoxic Activity of Compound I-8-e, I-9-e and I-10-f

In vitro cytotoxic activity of compound I-8-e, I-9-e and I-10-f weretested in HL-60 cells. As shown in Table 2, compound I-8-e and I-9-edemonstrated significant inhibition against HL-60 cancer cell lines.Compound I-8-e showed an IC₅₀ of 15 μM and compound I-9-e showed an IC₅₀of 5.8 μM against HL-60 cell. Compound I-9-e is an attractive candidatefor development as a novel anti-cancer agent.

TABLE 2 Compound IC₅₀ (μM) I-8-e 15 I-9-e 5.8 I-10-f >50 * HL-60 cellwere treated with compound I-8-e, I-9-e and I-10-f for 48 h. Aftertreatment, cells were harvested and examined using MTT assay. * IC₅₀value means the concentration causing 50% growth-inhibitory effect. *HL-60, leukemia cancer cell line.

What is claimed is:
 1. A phosphate derivative of 2-aryl-4-quinolonehaving the following formulas Ib or Ic:

wherein R₂′, R₃′, and R₄′ independently are H, (CH₂)_(n)CH₃,(CH₂)_(n)YH, Y(CH₂)_(n)CH₃, Y(CH₂)_(n)YH, Y(CH₂)_(n)NR₈R₉, X,(CH₂)_(n)NR₈R₉,

wherein n is an integer of 0-4, Y is O or S, X is F, Cl, or Br, and R₈and R₉ independently are H, (CH₂)_(n)YH,(CH₂)_(n)N(C_(n)H_(2n+1))(C_(m)H_(2m+1)) or (CH₂)_(n)CH₃, wherein n andY are defined as above, and m is an integer of 0-4; R₂, R₃, R₄ and R₅independently are H, (CH₂)_(n)CH₃, (CH₂)_(n)YH, Y(CH₂)_(n)CH₃,Y(CH₂)_(n)YH, Y(CH₂)_(n)NR₈R₉, X, (CH₂)_(n)NR₈R₉,

or R₃ and R₄ together is —Y(CH₂)_(n)Y—, wherein n, Y, X, R₈ and R₉ aredefined as above; and R₁ and R₁′ independently are H, L⁺, Na⁺, K⁺,N⁺R₈R₉R₁₀R₁₁ or benzyl wherein R₁₀ and R₁₁ independently are H,(CH₂)_(n)YH, (CH₂)_(n)N(C_(n)H_(2n+1))(C_(m)H_(2m+1)) or (CH₂)_(n)CH₃,n, m, R₈ and R₉ are defined as above.
 2. The phosphate derivativeaccording to claim 1, wherein R₂′, R₃′ and R₄′ are all H; or one of R₂′,R₃′ and R₄′ is F or OCH₃, and the others thereof are H.
 3. The phosphatederivative according to claim 1, wherein R₁ and R₁′ are benzyl.
 4. Thephosphate derivative according to claim 3, wherein R₂′, R₃′, R₄′, R₂ andR₅ are all H, and R₃ and R₄ together is —O(CH₂)O—.
 5. A pharmaceuticalcomposition for the killing of solid cancer cells, which comprises atherapeutically effective amount of a phosphate derivative of2-aryl-4-quinolone as set forth in claim 1 or a pharmaceuticallyacceptable salt thereof, as an active ingredient, in admixture with apharmaceutically acceptable carrier or diluent for the activeingredient, wherein the solid cancer cells comprise human breast cancer,wherein R₂, R₃, R₄, and R₅ are all H; or one of R₂, R₃, R₄ and R₅ is For OCH₃, and the others thereof are H; or R₂ and R₅ are H, and R₃ and R₄together is —O(CH₂)_(n)O—, wherein n is defined as in claim 1.